Among the glucose transporter (GLUT) family, GLUT4 is unique in that it is confined mainly to insulin-sensitive fat and muscle tissues and is the major insulin-responsive isoform. Insulin stimulation results in the recruitment of GLUT4 to the plasma membrane (PM), thereby allows increased glucose flux into the cells. Evidences indicate that irregularities in GLUT4 trafficking underlie obesity and type 2 diabetes. Elevated levels of sphingolipids (SLs) such as ceramide and ganglioside GM3 are typical in animal models of both diseases. SLs are an important class of lipid molecules that play roles in a wide variety of cell functions including cell-cell interaction, cell growth and differentiation and signal transduction. They interact with cholesterol to form membrane micordomains. The general hypothesis tested in this proposal is that GLUT4 trafficking is regulated by SL and cholesterol. As indicated in the Specific Aims, we will investigate the role of SLs and cholesterol in insulin-responsive GLUT4 translocation and internalization. The results from the proposed study will facilitate our understanding the GLUT4 trafficking and function in disease state and provide potential therapeutic approaches.